Abstract
Atomic force microscopy was applied to investigate the influence of protein and precipitant (sodium-potassium tartrate) concentration on thaumatin crystal growth mechanisms. At constant protein concentration, a decrease of salt concentration from 0.8 to 0.085 M caused a transition of the crystal growth mechanism from two-dimensional nucleation to dislocation growth. At different, fixed concentrations of salt, the protein concentration, which does not induce multiple crystal nucleation, was increased from 8 to 60 mg/ml with corresponding increases in the tangential velocity of growth steps from 5 to 17.5 nm/s. Results from these experiments suggest that a highly concentrated protein solution, as might be found in a protein rich phase, may not induce crystal nucleation, but can promote crystal growth if screw dislocations are present in the crystal.
Highlights
Unless seeding is employed, macromolecular crystals are always grown at very high levels of supersaturation necessary for nucleation, but supersaturation ranges non-ideal for growth
Crystals may fail to grow large because too little protein is available. This is primarily due to the fact that at high precipitant concentrations, crystals grow predominantly by the mechanism of two-dimensional nucleation, a mechanism that is sensitive to, and limited by protein concentration
We have further studied the growth of thaumatin crystals [10,11] under a variety of protein concentrations and salt concentrations using AFM
Summary
Macromolecular crystals are always grown at very high levels of supersaturation necessary for nucleation, but supersaturation ranges non-ideal for growth. Crystals may fail to grow large because too little protein is available This is primarily due to the fact that at high precipitant concentrations, crystals grow predominantly by the mechanism of two-dimensional nucleation, a mechanism that is sensitive to, and limited by protein concentration. This has been demonstrated for a number of macromolecular crystals by AFM [1,2,3,4,5,6,7,8,9]. This investigation, we believe, contributes to a better understanding of the considerations involved in choosing optimal crystallization conditions for macromolecules
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